Exfoliation syndrome is common and may affect 5-10% of Americans and up to 50% in some populations (i.e. Icelanders). As many as 10 to 50% of those with exfoliation syndrome develop high intraocular pressure and a secondary glaucoma - exfoliative glaucoma, which causes blindness and visual disability in millions worldwide. Exfoliative syndrome has a strong genetic basis, but complex inheritance. Genome-wide association studies (GWAS) of exfoliation syndrome have successfully identified 7 genetic factors that increase risk for disease: LOXL1, CACNA1A, POMP, TMEM136, AGPAT1, RBMS3, and SEMA6A. However, an animal model has not yet been built based upon these gene discoveries. There is a critical need for these human GWAS discoveries to be translated into animal models. We previously reported mice, B6-Lystbg-j, with a mutation in the Lyst gene, that have some features of exfoliation syndrome, but produce only tiny amounts of exfoliation material and do not develop glaucoma. We hypothesize that inactivating the 7 exfoliation syndrome risk factor genes using CRISPR/Cas9 methods will generate animal models of exfoliation syndrome and glaucoma. We further hypothesize that introducing these mutations on the B6-Lystbg-j background strain will take advantage of its subtle exfoliation syndrome features and facilitate developing a new line with pronounced features, high pressure and glaucoma. We have created CRISPR reagents to inactivate the seven exfoliation syndrome risk factors and we have begun injecting mouse embryos. We are injecting pools of CRISPR reagents for our target genes in order to get founder mice with one or more inactivated risk factor genes. All mutations are made in B6-Lystbg-j mice. AIM 1A: ASSESS FOUNDER MICE. We will assess the eyes of founder mice (in vivo) that have one or more inactivated exfoliation syndrome risk factor genes for exfoliation material. Those with exfoliative material in their eyes will be aged to 1 year, assessed for high intraocular pressure, and glaucomatous optic neuropathy. AIM 1B: GENERATE MICE WITH MULTIPLE INACTIVATED RISK FACTORS. We will conduct crosses to produce mice with multiple inactivated exfoliation syndrome risk factor genes on a B6-Lystbg-j background: 1) To produce mice with 2 heterozygously inactivated risk factor genes (with extreme individual phenotypes). 2) To produce mice with 2 risk factors (Loxl1, Cacn1a1) heterozygously inactivated (top GWAS hits). AIM 2A DETERMINE EXPRESSION PATTERNS OF RISK FACTOR GENES IN B6-Lystbg-j MICE. We will assess expression of the 7 risk factor genes at the mRNA and protein level in ocular tissues of B6-Lystbg-j mice relevant to exfoliation syndrome pathophysiology (lens, iris, and iridocorneal angle). AIM 2B DETERMINE EXPRESSION PATTERNS OF RISK FACTOR GENES IN OUR CRISPR/Cas9 MICE. INNOVATION. Pooling CRISPR guide RNAs to make mice with inactivated risk factor genes (alone and in combination). Creating the 1st animal model of exfoliation syndrome from GWAS hits as a step towards a cure.